K88 fimbriae of enterotoxigenic E. coli and their receptors in piglet small intestine
A study was designed to identify receptors for K88ac fimbriae of enterotoxigenic E.coli (ETEC) in the small intestinal mucosa of the piglet. K88ac fimbriae were purified and labelled via amino groups using digoxigenin-3-O-methylcarbonyl-Ε-aminocaproic acid-N-hydroxy-succinimide ester, and their binding activity was demonstrated in haemagglutination assay. Digoxigenin-labelled fimbriae were then used as probes in Western blot overlay analyses of piglet intestinal brush border membranes, and a family of high molecular weight glycoproteins (>250 kDa) that specifically bound K88ac fimbriae was detected. Expression of these glycoproteins was shown to correlate with susceptibility both to K88ac-mediated adhesion of E.coli in vitro and K88ac-mediated enterotoxicosis in vivo. A weakly adhesive piglet phenotype, identified in in vitro brush border cell adhesion assays, was found to be associated with the presence of a 200 kDa glycoprotein in the intestinal brush border membrane. It would seem that these glycoproteins are the receptors used by E.coli K88ac to adhere to the piglet intestinal epithelium, and their presence may be the basis for susceptibility to K88ac-mediated ETEC infections. In anticipation of being able to develop strategies for inhibiting pathogen adherence, the nature of the K88ac fimbrial-receptors was examined. Methods for the recovery of the receptor glycoproteins were assessed, and partial purification was achieved by affinity chromatography using K88ac fimbriae immobilised on Sepharose beads.